The Unfolded Protein Response, (UPR) is defined as the natural reaction of cells to stresses caused by the accumulation of misfolded/unfolded proteins within the endoplasmic reticulum (ER). UPR may cause: 1) degradation of mRNAs encoding secretory proteins, 2) inhibition of global protein synthesis, and 3) degradation of unfolded proteins that accumulate in the ER. While UPR works to restore ER homeostasis and promote cell survival, it changes the gene expression of stressed cells and affects their functions, and chronic (pathologic) UPR may cause various diseases. Non-syndromic dentinogenesis imperfecta (DGI)/dentin dysplasia (DD), a common inherited dentin disorder, is caused by mutations in one allele of the dentin sialophosphoprotein (DSPP) gene. Whether DGI/DD is caused by DSPP haploinsufficiency itself or by the accumulation of mutant DSPP in the ER is not understood. Preliminary in vitro data showed that 1) the secretion of mouse DSPP-P19L, a homolog of the human DSPP mutant, p. P17L, was impaired in the transfected cells, and 2) that the chemical chaperone, 4-phenylbutyrate (4-PBA), accelerated the secretion of mutant DSPP-P19L. Analyses with the recently created DsppP19L/+ knock-in mice showed that odontoblasts in DsppP19L/+ or DsppP19L/P19L mice failed to efficiently secrete mutant DSPP into the dentin matrix and that the tooth defects (chamber enlargement) of younger DsppP19L/+ and DsppP19L/P19L mice resembled human DGI Type III, whereas those at an older age (chamber obliteration) mimicked human DGI Type II. While DsppP19L/+ and DsppP19L/P19L odontoblasts had a markedly reduced level of DSPP mRNA, they showed an increased level of the phosphorylated form of inositol-requiring enzyme 1? (IRE1?), indicating the activation of one UPR branch. These findings lead to the central hypothesis that the mutant DSPP-P19L within the ER causes ER stress and pathologic UPR, resulting in DGI; alleviation of ER stress by facilitating the secretion of ER- retained DSPP-P19L may reduce or correct dentin defects. Three Aims are proposed to test this novel hypothesis: Aim 1 - To determine the pathological effects of intracellularly retained DSPP-P19L on odontoblasts and dentin formation, by a) examining if DSPP-P19L accumulates in the ER and causes ER dilation, b) analyzing the dentin defects, c) assessing the activities of all three UPR branches in the molars of DsppP19L/+ and DsppP19L/P19L mice, and d) determining if UPR associated with DSPP-P19L induces autophagy, apoptosis and pro-inflammatory responses. Aim 2 - To determine if chemical chaperones will relieve ER stress and prevent the dental defects of the DsppP19L/+ and DsppP19L/P19L mice, by treating these mice with 4-PBA. Aim 3 - To determine the roles of IRE1? in DSPP mRNA degradation and odontoblast differentiation and function, by a) assessing the influences of IRE1? activation and inactivation on DSPP mRNA, and b) analyzing the teeth of mice in which IRE1? is conditionally ablated. Completion of the proposed project will elucidate the molecular pathogenesis of DGI and help develop effective and non-invasive treatment modalities for DGI.